METTL14/m6A/CEBPD axis: A critical player in chronic cerebral hypoperfusion-associated neuroinflammation.

Abstract

Vascular dementia (VaD) is a major cause of cognitive decline in the elderly, with chronic cerebral hypoperfusion (CCH) as its core pathogeny linked to neuroinflammation. Microglial M1 polarization drives CCH-related neuroinflammation, but its epigenetic regulation remains unclear. N6-methyladenosine (m6A) modification, centered on methyltransferase-like 14 (METTL14), is abundant in the brain, while CCAAT enhancer binding protein delta (CEBPD) is associated with central nervous system inflammation. This study explored METTL14's role in microglial polarization and CCH pathology. In vitro, HMC3 cells induced by LPS/TNF-α (10 μg/mL, 50 ng/mL) showed upregulated METTL14 and CEBPD. METTL14 knockdown via short hairpin RNA inhibited the expression of M1 phenotypic markers (iNOS, CD86), reduced the release of pro-inflammatory factors (IL-1β, IL-6), and blocked NF-κB nuclear translocation, indicating that METTL14 silencing suppresses LPS/TNF-α-induced microglial M1 polarization and inflammatory response. In bilateral common carotid artery occlusion (BCCAo)-induced CCH rats, hippocampal METTL14/CEBPD was elevated. Adeno-associated virus-mediated METTL14 knockdown not only attenuated neuroinflammation and neuronal apoptosis in hippocampal CA1, but also significantly improved spatial learning memory and object recognition ability. Mechanistically, RIP-PCR and MeRIP-qPCR confirmed METTL14 enhances CEBPD mRNA stability via m6A modification. CEBPD overexpression reversed METTL14 knockdown's effect, identifying it as a key downstream mediator. In conclusion, METTL14 knockdown impairs CEBPD mRNA stability via m6A, inhibits TLR4/MyD88/NF-κB pathway and microglial M1 polarization, alleviating CCH-related neuroinflammation and cognitive impairment, providing a potential epigenetic target for VaD.